Modulation of antioxidant enzymes,reactive oxygen species,and glutathione levels in manganese superoxide dismutase-overexpressing NIH/3T3 fibroblasts during the cell cycle

NIH/3T3 mouse embryo fibroblasts were transfected with the cDNA for manganese superoxide dismutase (MnSOD). Previous studies showed characteristic unique AE profiles in nonsynchronous populations of parental, control plasmid-transfected, and MnSOD-overexpressing NIH/3T3 cell lines. However, the present study showed that during S and M phases of the cell cycle, antioxidant enzyme (AE) levels were altered in MnSOD-overexpressing cell lines towards levels in S and M phases of parental and control plasmid-transfected cells. Because of the demonstration that MnSOD overexpression inhibits cell growth in both nonmalignant and malignant cells, the present study was designed to measure AEs, reactive oxygen species (ROS), and glutathione levels in various phases of the cell cycle in both parental NIH/3T3 cells and NIH/3T3 cells overexpressing MnSOD, to try to determine whether AEs, ROS, and glutathione levels could have a possible regulatory role in cell cycle progression. In all cell lines studied, ROS levels were lower in M than S phase of the cell cycle. Total glutathione and glutathione disulfide levels were greatly increased during the M phase of the cell cycle compared with quiescence and S phase in all cell lines studied. This suggests that oxidative stress exists in M phase of the cell cycle with total glutathione levels increased to decrease oxidative stress. Analysis of MnSOD-overexpressing cell clones showed a correlation of decreased cell growth with an increase in ROS in S phase of the cell cycle and a decrease in glutathione in mitosis. The data strongly suggest that specific levels of cell redox state are necessary for cells to successfully progress through the various phases of the cell cycle. J. Cell. Physiol. 177:148–160, 1998. © 1998 Wiley-Liss, Inc.